In this study, the constant-region genes (C alpha, C beta and C gamma) that encode the T-cell antigen receptor (TCR) alpha, beta and gamma chains were cloned from mandarin fish, Siniperca chuatsi Basilewsky, an important freshwater fish species in China. The complementary DNA sequences of C alpha, C beta and C gamma were 843, 716 and 906base pairs (bp) in length and had a 465-, 289- and 360-bp 3 ' untranslated region, encoding 125, 142 and 182 amino acids, respectively. The amino-acid sequences of the constant regions of mandarin fish TCR alpha, beta and gamma chains (encoded by C alpha, C beta and C gamma, respectively) were most similar to those of their teleost counterparts, showing 60% similarity with pufferfish, 48% similarity with Atlantic salmon and 57% similarity with flounder, respectively. The phylogenetic analysis revealed that the mandarin fish C alpha, C beta and C gamma were clustered, respectively, with their vertebrate counterparts. The mandarin fish C alpha, C beta and C gamma could also be separated into four domains: immunoglobulin; connecting peptide (CP); transmembrane (TM); and cytoplasmic tail. Several conserved features in mammalian TCRs were also found in those of mandarin fish, such as a conserved cysteine residue in the CP domain of C alpha, necessary for creating an interchain disulphide bond with the TCR beta chain, and a conserved antigen receptor TM motif in C alpha and C beta. Meanwhile, transcripts of C alpha, C beta and C gamma were detectable in all examined organs, with a stronger signal observed in lymphoid organs. In addition, the temporal transcriptional changes for C alpha and C gamma were investigated, 1, 2, 3, 4, 5, 6 and 8weeks after stimulation with Flavobacterium columnare, in head kidney, spleen, blood, thymus, gill and intestine, using real-time polymerase chain reaction. The results demonstrated stimulation-dependent up-regulations in almost all tissues examined, which indicates that T cells may play important roles in preventing mandarin fish from bacterial invasion. In particular, apart from thymus, T cells were distributed mainly in gill and intestine, where striking up-regulation of C gamma was also observed. These results will facilitate functional studies of teleost TCRs and T cells.

英文摘要:

In this study, the constant-region genes (C alpha, C beta and C gamma) that encode the T-cell antigen receptor (TCR) alpha, beta and gamma chains were cloned from mandarin fish, Siniperca chuatsi Basilewsky, an important freshwater fish species in China. The complementary DNA sequences of C alpha, C beta and C gamma were 843, 716 and 906base pairs (bp) in length and had a 465-, 289- and 360-bp 3 ' untranslated region, encoding 125, 142 and 182 amino acids, respectively. The amino-acid sequences of the constant regions of mandarin fish TCR alpha, beta and gamma chains (encoded by C alpha, C beta and C gamma, respectively) were most similar to those of their teleost counterparts, showing 60% similarity with pufferfish, 48% similarity with Atlantic salmon and 57% similarity with flounder, respectively. The phylogenetic analysis revealed that the mandarin fish C alpha, C beta and C gamma were clustered, respectively, with their vertebrate counterparts. The mandarin fish C alpha, C beta and C gamma could also be separated into four domains: immunoglobulin; connecting peptide (CP); transmembrane (TM); and cytoplasmic tail. Several conserved features in mammalian TCRs were also found in those of mandarin fish, such as a conserved cysteine residue in the CP domain of C alpha, necessary for creating an interchain disulphide bond with the TCR beta chain, and a conserved antigen receptor TM motif in C alpha and C beta. Meanwhile, transcripts of C alpha, C beta and C gamma were detectable in all examined organs, with a stronger signal observed in lymphoid organs. In addition, the temporal transcriptional changes for C alpha and C gamma were investigated, 1, 2, 3, 4, 5, 6 and 8weeks after stimulation with Flavobacterium columnare, in head kidney, spleen, blood, thymus, gill and intestine, using real-time polymerase chain reaction. The results demonstrated stimulation-dependent up-regulations in almost all tissues examined, which indicates that T cells may play important roles in preventing mandarin fish from bacterial invasion. In particular, apart from thymus, T cells were distributed mainly in gill and intestine, where striking up-regulation of C gamma was also observed. These results will facilitate functional studies of teleost TCRs and T cells.